Di- or tripeptide renin inhibitors containing lactam conformational restriction in ACHPA

ABSTRACT

Enzyme di- or tripeptides of the formula: ##STR1## and analogs thereof which inhibit renin and are useful for treating various forms of renin-associated hypertension, hyperaldosteronism and congestive heart failure; compositions containing these renin-inhibitory peptides, optionally with other antihypertensive agents; and methods of treating hypertension, hyperaldosteronism or congestive heart failure or of establishing renin as a causative factor in these problems which employ these novel peptides.

This is a continuation, of application Ser. No. 403,986, filed Sept. 6,1989 now abandoned which is a continuation of U.S.S.N. 298,815 filed1/18/89 now abandoned which was a continuation of U.S.S.N. 108,344 filed10/14/87 and now abandoned.

The present invention is concerned with novel di or tripeptides whichinhibit the angiotensinogen cleaving action of the proteolytic enzyme,renin, with pharmaceutical compositions containing the novel peptides ofthe present invention as active ingredients, with methods of treatingrenin-associated hypertension, hyperaldosteronism, and congestive heartfailure, with diagnostic methods which utilize the novel peptides of thepresent invention, and with methods of preparing the novel peptides ofthe present invention.

BACKGROUND OF THE INVENTION

Renin is an endopeptidase (molecular weight about 40,000) produced andsecreted by the juxtaglomerular cells of the kidney. Renin has a highspecificity for and cleaves the naturally-occurring plasma glycoprotein,angiotensinogen, at only the 10, 11 peptide bond, i.e., between the 10th(Leu) and 11th (Leu) amino acid residues in the equine substrate, asdescribed by Skeggs et al, J. Exper. Med. 1957, 106, 439, or between Leu10 and Val 11 in the human renin substrate, as elucidated by Tewksburyet al., Circulation 59, 60, Supp. II: 132, Oct. 1979.

This cleavage of its tetradecapeptide substrate, angiotensinogen, byrenin splits off the decapeptide, angiotensin I, which is thought to behemodynamically-inactive, but which is converted in the lungs, kidney orother tissue by angiotensinconverting enzyme (ACE) to the potent pressoroctapeptide, angiotensin II. Angiotensin II then causes constriction ofthe arterioles and is also believed to stimulate release of thesodium-retaining hormone, aldosterone, from the adrenal gland, therebycausing a rise in extra cellular fluid volume. Thus, therenin-angiotensin system plays an important role in normalcardiovascular homeostasis and in some forms of elevated blood pressure(hypertension).

Inhibitors of angiotensin I converting enzyme have proven useful in themodulation of the renin angiotensin system. Consequently, specificinhibitors of the catalytic and rate-limiting enzymatic step thatultimately regulates angiotensin II production, the action of renin onits substrate, have also been sought as effective investigative tools,as well as therapeutic agents in the treatment of hypertension andcongestive heart failure.

Renin antibody, pepstatin (another aspartic proteinase, like renin),phospholipids, and substrate analogs, including tetrapeptides and octa-to tridecapeptides, with inhibition constants (K_(i)) in the 10⁻⁻³ 10⁻⁻⁶M region, have been studied.

Umezawa et al., in J. Antibiot. (Tokyo) 23: 259-262, 1970, reported theisolation of a peptide, pepstatin, from actinomyces that was aninhibitor of aspartyl proteases such as pepsin, cathepsin D, and renin.Gross et al., Science 175:656, 1972, reported that pepstatin reducesblood pressure in vivo after the injection of hog renin intonephrectomized rats. However, pepstatin has not found very wideapplication as an experimental agent because of its limited solubilityand its inhibition of a variety of other acid proteases in addition torenin.

Many efforts have been made to prepare a specific renin inhibitor basedon pig renin substrate analogy, since such analogy has been shown tocorrelate well with and predict human renin inhibitor activity. Theoctapeptide amino acid sequence extending from histidine-6 throughtyrosine 13 ##STR2## has been shown to have kinetic parametersessentially the same as those of the full tetradecapeptide reninsubstrate.

Kokubu et al., Biochem. Pharmacol., 22, 3217-3223, 1973, synthesized anumber of analogs of the tetrapeptide found between residues 10 to 13,but while inhibition could be shown, inhibitory constants were only ofthe order of 10⁻⁻³ M. Analogs of a larger segment of renin substratehave been also synthesized, e.g., Burton et al., Biochemistry 14:892-3898, 1975, and Poulsen et al., Biochemistry 12: 3882, 1973, but alack of solubility and weak binding (large inhibitory constant)generally resulted.

Modifications to increase solubility soon established that theinhibitory properties of the peptides are markedly dependent on thehydrophobicity of various amino acid residues. These modifications alsoestablished that increasing solubility by replacing lipophilic aminoacids with hydrophilic isosteric residues can become counter-productive.Other approaches to increasing solubility have also had limited success.

Modifications designed to increase binding to renin have also been made,but here too, with mixed results.

A series of inhibitors of renin have been disclosed which contain theunnatural amino acid, statine: see, e.g., Veber et al, U.S. Pat. Nos.4,384,994 and 4,478,826; Evans et al, U.S. Pat. No. 4,397,786; Boger etal, Nature, 1983, 303, 81-84 and U.S. Pat. Nos. 4,470,971; 4,485,099;4,663,310 and 4,668,770; Matsueda et al, EP-A 128 762, 152 255; Morisawaet al., EP-A 186 977; Riniker et al, EP-A 111 266; Bindra et al, EP-A155 809; Stein et al, Fed. Proc. 1986, 45, 869; and Holzemann et al,German Offenlegungsschrift DE 3438545. Attempting to explain the effectof statine, Powers et al., in Acid Proteases, Structure, Function andBiology, Plenum Press, 1977, 141-157, observed that in pepstatin,statine occupies the space of the two amino acids on either side of thecleavage site of a pepsin substrate and Tang et al., in Trends inBiochem. Sci., 1:205-208 (1976) and J. Biol. Chem., 251:7088-94, 1976,pointed out that the statine residue of pepstatin resembles thetransition state for pepsin hydrolysis of peptide bonds.

Renin inhibitors containing other peptide bond isosteres, including areduced carbonyl isostere have been disclosed by M. Szelke et al, inwork described in published European Patent Applications 45 665 and 104041; in U.S. Pat. No. 4,424,207, and in PCT Int. Appl. WO 84/03044; inNature, 299, 555 (1982); Hypertension, 4, Supp. 2, 59, 1981; and BritishPatent 1,587,809. In Peptides, Structure and Function: Proceedings ofthe Eighth American Peptide Symposium, ed. V. J. Hruby and D. H. Rich,p. 579, Pierce Chemical Co., Rockford, IL., 1983, Szelke et al alsoshowed isosteric substitutions at the Leu-Leu site of cleavage,resulting in compounds with excellent potency.

Other peptide bond isosteres have then been disclosed in Buhlmayer et alin EP-A 144 290 and 184 550; Hester et al, EP-A 173 481; Raddatz, EP-A161 588; Dann et al, Biochem. Biophys Res. Commun. 1986, 134, 71-77;Fuhrer et al, EP-A 143 746; Kamijo et al, EP-A 181 110; Thaisrivongs etal, J. Med. Chem., 1985, 28, 1553-1555; Ryono et al., EP A 181 071; andEvans et al, U.S. Pat. No. 4,609,641.

Other modifications which have been tried include preparing renininhibitors with non peptide C-termini, such as disclosed in EuropeanPublished Applications 172 346 and 172 347; Evans et al, J. Med. Chem.,1985, 28, 1755-1756; Bock et al, Peptides, Structure and Function:Proceedings of the Ninth American Peptide Symposium, ed. C. M. Deber etal, pp.751-754, Pierce Chemical Co., Rockford, IL, 1985; and Plattner etal, in Abstracts from the 191st National Meeting of the AnericanChemical Society, April, 1986. Kokubu et al, in Hypertension, 1985, 7,Suppl. I, p. 8-10 and Matsueda et al, in Chemistry Letters, 1985,1041-1044 and in European Published Applications 128 762 and 152 255disclosed peptide aldehyde renin inhibitors, and Hanson et al inBiochem. Biophys. Res. Commun. 1985, 132, 155-161, reported peptideglycol inhibitors.

These various renin inhibitors all generally comprise peptide-basedinhibitors in which a sequence of the type:...A--B--D--E--F--G--J--K--L... , where G is a peptide bond mimic andA,B,D,E,F,J,K, and L may individually be absent or may representnaturally-occurring or modified amino acids. Typical sequences of thistype include: ##STR3## where the N-terminus typically comprises an aminoacid protecting group such as BOC or CBZ, and the N-terminal amino acidsare Pro--Phe--His or Phe--His.

Lower molecular weight renin-inhibitory di-or tripeptides comprisingacyclic 2-substituted-4-amino-5 cycl-ohexyl-3 hydroxy-pentanoic acid(ACHPA) have been disclosed in U.S. Pat. application 45,941, filed May4, 1987, and other lower molecular weight peptides have been disclosedin Sham, EP 184 855, Bindra et al, EP 155 809, and Matsueda et al, EP152 255.

It was an object of this invention to prepare lower molecular weightpeptides which have enhanced biological potency in inhibiting the reninenzyme. It was also an object to prepare shortened peptide sequenceswhich incorporate at the C terminus a stabilizing, conformationallyconstrained dipeptide mimic to replace the 10- and 11-position aminoacids in the analogous natural substrate. It was a further object toinclude strategically-located substituents at the C- and/or N-terminiiof a shortened peptide which confer increased potency whileconstructively altering the physical properties of these peptides. Itwas an additional object of this invention to prepare peptides whichhave greater oral bioavailability and increased duration of action. Itwas still a further object of this invention to prepare novel peptideswhich are more useful antihypertensive agents, and compounds useful intreating hyperaldosteronism and congestive heart failure.

DETAILED DESCRIPTION OF THE INVENTION

The present invention is directed to renin-inhibitory di and tripeptidesof the structure: ##STR4## wherein: A is hydrogen; C₁ -C₆ -alkyl; aryl,where aryl is unsubstituted or mono-, di or trisubstituted phenyl,wherein the substituent(s) is/are independently selected from the groupconsisting of C₁ -C₇ -alkyl, amino, mono- or di-C₁ -C₄ -alkylamino,amino-C₁ -C₄ -alkyl, hydroxy-C₁ -C₄ -alkyl, phenyl-C₁ -C₄ -alkyl, mono-or di-C₁ -C₄ aklylamino-C₁ -C₄ -alkyl, guanidyl, guanidylC₁ -C₄ - alkyl,hydroxyl, C₁ -C₄ -alkoxy, trifluoromethyl, halo, CHO, --CO₂ H, --CONH₂,--CONH-C₁ -C₄ -alkyl, --CON(C₁ -C₄ -alkyl)₂, --CO-C₁ -C₄ -alkyl,--(CH₂)hd m-13 ⁺ N(R³)₂ R⁴ A.sup.⊖, where R³ is C₁ -C₄ alkyl, --(CH₂)₄-, --(CH₂)₅ --or --(CH₂)₂ -O-(CH₂)₂ --;R⁴ is C₁ -C₄ -alkyl, C₁ -C₄-hydroxyalkyl, C₁ - C₄ -carboxyalkyl, or --CH₂ -phenyl; A.sup.⊖ is acounterion selected from the group consisting of singlenegatively-charged ions, such as chloride, bromide, perchlorate,benzoate, benzene sulfonate, tartrate, maleate, hemitartrate, andacetate; and m is 0-to-3; --CO₂ -C₁ -C₄ -alkyl, --CO₂ -C₁ -C₄ -alkoxy-C₂-C₄ --alkyl, ##STR5## where A.sup.⊖ and m are as defined above, and--NR⁵ R⁶, where R⁵ and R⁶ are independently hydrogen, unsubstituted ormonosubstituted C₁ -C₄ -alkyl, wherein the substituent is amino, mono-or di--C₁ -C₄ --alkylamino or .sup.⊕ N(R³)₂ R⁴ A.sup.⊖, where R³, R⁴ andare as defined above; Het, where Het is an unsubstituted or mono-ordisubstituted 5- or 6-membered mono or bicyclic or benzofused 5 or 6membered heterocyclic ring, where the one or two heteroatoms areindependently selected from the group consisting of N, O, S, NO, SO, SO₂or quaternized N, and the substituent(s) is/are independently selectedfrom the group consisting of hydroxyl, thiol, C₁ -C₆ -alkyl, CF₃, C₁ -C₄-alkoxy, halo, aryl, as defined above, aryl-C₁ -C₄ -alkyl, amino mono-or di C₁ -C₄ -alkylamino, amino-C₁ -C₄ -alkyl, hydroxy-C₁ -C₄ -alkyl,mono or di-C₁ -C₄ -alkylamino-C₁ -C₄ -alkyl, guanidyl, guanidylC₁ -C₄-alkyl, CHO, CO₂ H, CO₂ -C₁ -C₃ -alkyl, CONH₂, CONH-C₁ -C₄ -alkyl,CON(C₁ -C₄ -alkyl)₂, NR⁵ R⁶, ##STR6## and -(CH₂)_(m) --.sup.⊕ N(R³)₂ R⁴A.sup.⊖, wherein R⁵, R⁶, A.sup.⊖, m, R³ and R⁴ are as defined above, orwhen the heteroatom is N, the substituents are alternatively (CH₂)q or--(CH₂)₂ --O--(CH₂)₂ - and form a ring with the N-atom, wherein q is3-to-6; where R² is C₁ -C₇ -alkyl; hydrogen; Het, as defined above;aryl, as defined above; mono substituted C₁ -C₅ -alkyl, wherein thesubstituent is selected from the group consisting of aryl, as definedabove; Het, as defined above; hydroxyl; -CO₂ H; CO₂ R⁷, where R⁷ is C₁-C₅ -alkyl, aryl, as defined above, and aryl-C₁ -C₄ -alkyl; CONH₂ ;--CONH-R⁷ or --S(O)_(n) --R⁷, wherein n is 0-to-2 and R⁷ is as definedabove; C₁ -C₄ -alkoxy; C₃ -C₇ -cycloalkyl; amino; mono- or di--C₁ -C₄-alkylamino; NH-aryl, --NH-CH₂ -aryl or --CO-aryl, where aryl is asdefined above; and --NH-Het, --NH-CH₂ -Het or --CO-Het, where Het is asdefined above; ##STR7## where R⁷ is as defined above; or ##STR8## whereR⁹ is C₁ -C₅ -alkyl, aryl, as defined above, or Het, as defined above;

B and D are independently ##STR9## where R¹² is hydrogen, C₁ -C₅ -alkylor CH₂ -aryl, wherein aryl is as defined above; and R² is as definedabove; ##STR10## where R² is as defined above; or either B or D, but notboth simultaneously, is absent;

R¹ is hydrogen; C₃ -C₆ -alkyl; aryl, as defined above; unsubstituted,mono-, di- or trisubstituted C₃ -C₇ -cycloalkyl, where thesubstituent(s) is/are selected from the group consisting of C₁ -C₄-alkyl, trifluoromethyl, hydroxyl, C₁ -C₄ -alkoxy and halo; orunsubstituted or 4-monosubstituted 1,3-dithiolan-2-yl or unsubstitutedor 4-mono-substituted 1,3-dithian-2-yl, where the substituent is(CH₂)_(m) aryl, where m and aryl are as defined above; ##STR11## whereR¹³ and R¹⁴ are independently hydrogen; C₁ -C₇ -alkyl; C₂ -C₇ -alkenyl;-CO₂ H; -CONH₂ ; CO₂ R⁷, --CO--NH--R⁷ or --CO--N(R⁷) , wherein R⁷ is asdefined above; mono-substituted C₁ -C₅ -alkyl, wherein the substituentis selected from the group consisting of azido; halo; hydroxy; C₁ -C₅-alkoxy; aryl, aryl--CH₂ O, aryloxy, aryl--COO--, aryl--CH₂ --NH--orarylamino, where aryl is as defined above; C₁ -C₅ -alkyl--CO₂ --; R⁷NH--COO--, R⁷ --CO--NH--, R⁷ --NH--CO--NH--or R⁷ --S(O)_(n), where n andR⁷ are as defined above; amino; mono or di-C₁ -C₄ -alkylamino; and Het,as defined above; or R¹³ and R¹⁴ are connected to form a polymethylenechain of the formula, --(CH₂)_(p), where p is 2 to 6; or ##STR12## whereR¹³ and R¹⁴ are as defined above; Y is CH₂, O, S, SO or SO₂ ; or

Y--X is --(CH₂)₄ --; and

E is hydrogen; aryl, as defined above; Het, as defined above; C₂ -C₇-alkenyl; or unsubstituted or mono- substituted C₁ -C₇ -alkyl orunsubstituted or mono-substituted C₃ -C₇ -cycloalkyl, where thesubstituent is selected from the group consisting of aryl, --CO aryl,--NH--aryl or --O--aryl, wherein aryl is as defined above; Het,--NH--Het, --O--Het, --CO--Het, --NH--CO--Het, CO--NH--Het,--CO--NH--CH₂ --Het or --O--CO--Het, wherein Het is as defined above;azido; C₃ -C₇ -cycloalkyl; halo; hydroxyl; C₁ -C₄ -alkoxy; --COOH;--O--CO--R⁷, --O--CO--NH R⁷, --NH--CO--R⁷, --NH--CO--NH R⁷, --S(O)_(n)--R⁷, --CO₂ R⁷ or --CO--NH--R⁷, wherein R⁷ and n are as defined above;amino; mono- or di C₁ -C₄ -alkylamino; CHO; and --.sup.⊕ N(R³)₂ R⁸A.sup.⊖, ##STR13## where R⁸ is C₁ -C₄ -alkyl, C₁ -C₄ hydroxyalkyl, C₁-C₄ -carboxyalkyl, --CH₂ -aryl, wherein aryl is as defined above, or--CH₂ --Het, wherein Het is as defined above, and R³ and A.sup.⊖ are asdefined above;

and pharmaceutically acceptable salts thereof.

In the peptides of the present invention, the components havingasymmetric centers occur as racemates, racemic mixtures and asindividual diastereomers, with all isomeric forms generally beingincluded in the present invention. In particular, asymmetric carbonatoms at the 2, 3 and 4 positions in peptides of Formula I preferablyhave an S configuration.

When any variable (e.g., aryl, Het, m, n, R², R³, R⁷, A⁻, etc.) occursmore than one time in any variable or in formula I, its definition oneach ocurrence is independent of its definition at every otheroccurrence.

As used herein, "alkyl" is intended to include both branched- andstraight chain saturated aliphatic hydrocarbon groups having thespecified number of carbon atoms (Me is methyl, Et is ethyl); "alkoxy"represents an alkyl group of indicated number of carbon atoms attachedthrough an oxygen bridge; and "C₃ -C₇ -cycloalkyl" is intended toinclude saturated ring groups, such as cyclopropyl, cyclobutyl,cyclopentyl, cyclohexyl and cycloheptyl. "Alkanoyl" is intended toinclude those alkylcarbonyl groups of specified number of carbon atoms,which are exemplified by formyl, acetyl, propanoyl and butanoyl;"alkenyl" is intended to include hydrocarbon chains of either a straightor branched- configuration and one unsaturation, which may occur at anypoint along the chain, such as ethenyl, propenyl, butenyl, pentenyl, andthe like, and includes E and Z forms, where applicable; and "arylalkyl"represents aryl groups as herein defined which are attached through astraight- or branched- chain alkyl group of specified number of carbonatoms, such as, for example, benzyl, phenethyl, 3,3-diphenylpropyl,3-indolymethyl, and the like. "Halo", as used herein, means fluoro,chloro, bromo and iodo, and "counterion" is used to represent a small,single negatively-charged specie, such as chloride, bromide, hydroxide,nitrate, acetate, benzoate, perchlorate, benzene sulfonate, tartrate,hemitartrate, maleate, and the like.

As used herein, with exceptions as noted, "aryl" is intended to meanphenyl (Ph), which is optionally-substituted by from one- to three-members independently selected from the group consisting of C₁ -C₇-alkyl, amino (Am), mono- or di-C₁ -C₄ -alkylamino, phenyl C₁ -C₄-alkyl, amino-C₁ -C₄ alkyl, hydroxy-C₁ -C₄ -alkyl, mono or di-C₁ -C₄-alkylamino-C₁ -C₄ -alkyl, hydroxyl, guanidyl, guanidyl-C₁ -C₄ -alkyl,C₁ -C₄ -alkoxy, CF₃, halo, CHO, CO₂ H, CONH₂, CONH--C₁ -C₄ -alkyl,CON(C₁ -C₄)₂, CO--C₁ -C₄ -alkyl or (CH₂)_(m) --.sup.⊕ N(R³)₂ R⁴ A.sup.⊖,wherein R³, R⁴ and m are as defined above and A.sup.⊖ is counterion, asdefined herein. "Aroyl" is intended to include those aryl carbonylgroups which are exemplified by phenoyl.

The term "Het", as used herein, represents a 5- to 7-membered mono- orbicyclic heterocyclic ring which is either saturated or unsaturated, andwhich consists of carbon atoms and one or two heteroatoms selected fromthe group consisting of N, O and S, and wherein the nitrogen and sulfurheteroatoms may optionally be oxidized, and the nitrogen heteroatom mayoptionally be quaternized, and including any bicyclic group in which anyof the above defined heterocyclic rings is fused to a benzene ring.Heterocycles which contain nitrogen are preferred. In the case of aheterocyclic ring containing one or more nitrogen atoms, the point ofattachment may be at one of the nitrogen atoms, or at any carbon atom.Examples of such heterocyclic elements include piperidyl, piperidinyl,piperazinyl, 2-oxopiperazinyl, 2 oxopyrolodinal, 2-oxopiperidinyl,2-oxoazepinyl, azepinyl, pyrryl, pyrrolinyl, 4-piperidonyl,pyrrolidinyl, pyrazolyl, pyrazolinyl, pyrazolidinyl, imidazolyl,imidazolinyl, imidazolidinyl, pyridyl, pyrazinyl, pyrimidinyl,pyridazinyl, oxazolyl, oxazolidinyl, isoxazolyl, isoxazolidinyl,morpholinyl, thiazolyl, thiazolidinyl, isothiazolyl, quinucli-dinyl,isothiazolidinyl, indolyl, guinolinyl, isoguinolinyl, benzimidazolyl,benzothiazolyl, benzoxazolyl, furyl, thienyl, benzothienyl,thiamorpholinyl, thiamorpholinyl sulfoxide, and thiamorpholinyl sulfone.The heterocyclic moiety is further optionally-substituted by from one tofour- members independently selected from the group consisting ofhydroxyl, thiol, C₁ -C₆ -alkyl, CF₃, C₁ -C₄ -alkoxy, halo, aryl, aryl-C₁-C₄ -alkyl, amino, mono- or di-C₁ -C₄ -alkylamino, amino-C₁ -C₄-alkylamino, amino-C₁ -C₄ -alkyl, hydroxy-C₁ -C₄ -alkyl, di-C₁ -C₄-alkylamino-C₁ -C₄ -alkyl, guanidyl, guanidyl C₁ -C₄ -alkyl, CHO, CO₂ H,CO₂ -C₁ -C₄ -alkyl, CONH₂, CONH-C₁ -C₄ -alkyl, CON(C₁ -C₄ -alkyl)₂,--NR⁵ R⁶, ##STR14## or --(CH₂)_(m) --.sup.⊕ N(R³)₂ R⁸ A.sup.⊖,

wherein R⁵, R⁶, A.sup.⊖, m, R³ and R⁸ are as defined above, or when theheteroatom is N, the substituents on the N-atom are -13 (CH₂)_(q) --or--(CH₂)₂ --O--(CH₂)₂ --and form a ring with the N-atom, wherein q is asdefined above.

The following additional abbreviations have also been used herein:

    ______________________________________    Abbreviated    Designation   Amino Acid/Residue    ______________________________________    ACHPA         (3S,4S)-4-amino-5-cyclohexyl-3-                  hydroxypentanoic acid    Ala           L-alanine    Arg           L-arginine    Cys           cysteine    Gly           L-glycine    His           D- or L-histidine    HomoPhe       homologated phenylalanine    HomoTrp       homologated tryptophan    HomoTyr       homologated tyrosine    Ile           L-isoleucine    Leu           L-leucine    Lys           L-lysine    Met           L-methionine    Nle           norleucine    Nva           norvaline    Orn           L-ornithine    (p-MeO)Phe    L-para-methoxyphenylalanine    Phe           L-phenylalanine    Pro           proline    Sar           L-sarcosine (N-methylglycine)    Ser           L-serine    Sta           statine    Thr           L-threonine    Trp           L-tryptophan    Tyr           L-tyrosine    Val           L-valine                  Protecting Group    BOC            .sub.- t-butyloxycarbonyl    CBZ           benzyloxycarbonyl(carbobenzoxy)    DNP           2,4-dinitrophenyl    IPOC          isopropoxycarbonyl                  Activating Group    HBT(HOBt)     1-hydroxybenzotriazole hydrate    HOSU          N-hydroxysuccinimide                  Condensing Agent    DCCI (DCC)    dicyclohexylcarbodiimide    DPPA          diphenylphosphorylazide    EDC           1-(3-dimethylaminopropyl)-3-ethyl                  carbodiimide hydrochloride                  Reagent    (BOC).sub.20  di- .sub.- t-butyl dicarbonate    DIBAL         diisobutylaluminum hydride    DIPEA         diisopropylethylamine    DMAP          4-(dimethylamino)pyridine    TEA           triethylamine    TFA           trifluoroacetic acid    LAH           lithium aluminum hydride    LDA           lithium diisopropylamide    MCPBA         3-chloroperoxybenzoic acid    NMM           N-methyl morpholine    PPTS          pyridinium para-toluenesulfonate    TBAF          tetra- -n-butylammonium fluoride                  Solvent    HOAc (AcOH)   acetic acid    DMF           dimethylformamide    DMSO          dimethyl sulfoxide    EtOAc         ethyl acetate    EtOH          ethanol    Et.sub.2 O    ether    MeOH          methanol    THF           tetrahydrofuran    ______________________________________

The novel renin inhibitory peptides of the present invention may begeneralized and alternately described in terms of common amino acidcomponents and closely-related analogs thereof, in accordance withformula I, wherein A, R¹, X and E are as defined under Formula I;

B is Absent, Ala, Leu, Phe, HomoPhe, (p--MeO)Phe, Tyr, Trp, HomoTrp or##STR15## where R² is as defined above; and D is Absent, Ala, Ser, Met,Thr, Phe, Tyr, Trp, His, Lys, Orn, Arg or Val, such that B and D are notsimultaneously absent.

In terms of substrate analogy, a unique aspect and essential feature ofthe present invention is the substitution of the ##STR16## component forthe double amino acid sequence, Leu¹⁰ -Val¹¹ in the endogenous humanrenin substrate ##STR17## which substitution for both amino acids at thecleavage site rather than just one is believed to result in an improvedsubstrate analogy. This invention's peptides particularly comprise novellactam versions of rigid 4-amino-5-cyclohexyl-3-hydroxy-pentanoic acid(ACHPA), which enables stereo-specific placement of substituents, andallows the ##STR18## component to better mimic the Leu¹⁰ -Val¹¹dipeptide moiety in the natural substrate.

It will be understood that closely-related analogs of the above commonamino acids, for example, aliphatic amino acids in addition to Ala, Val,Leu, and Ile, such as α-aminobutyric acid (Abu), substituted phenylderivatives of Phe, and N.sup.α -methyl amino acids, are included in thebroad description of the novel inhibitory peptides of the presentinvention represented by Formula I and related definitions.

Preferred renin-inhibitory peptides are those wherein A is R² --CO--, R⁹--SO₂ --, C₁ -C₄ -alkyl-O-CO--or R⁷ --NH--CO--, wherein R², R⁷ and R⁹are as defined above; B is absent (when D is present), L-phenylalanyl orderivatives thereof substituted on the aromatic ring by para-methoxy or##STR19## D is absent (when B is present), L-histidyl or L valinyl; R¹is cyclohexyl; Y is O or CH₂ ; R¹³ and R¹⁴ in either definition of X aresimultaneously or independently hydrogen or methyl; and E is C₁ -C₆-alkyl, --(CH₂)_(r) --.sup.⊕ N(R³)₂ R⁸ CH₃ CO.sup.⊖₂, wherein r is 2 or3 and R³ and R⁸ are as defined above, ##STR20## wherein R⁸ is as definedabove. The preferred stereochemistry at the 2,3 and 4 positions is S.

Representative preferred renin-inhibitory peptides of the presentinvention include the following compounds having the structure:##STR21## wherein in the structures:

    __________________________________________________________________________    A            B             D  Y    X        E    __________________________________________________________________________     ○1       Boc       Phe           His                                  CH.sub.2                                       CH.sub.2 CH.sub.2                                                n-Bu       (CH.sub.3).sub.2 CHSO.sub.2                 "             "  "    "        "        ##STR22##                 "             "  "    "        "       (CH.sub.3).sub.2 CHSO.sub.2                 CH.sub.2 CH(CH.sub.2 Ph)CO                               "  "    "        "     ○5       --                  ##STR23##    "  "    "        "       Boc       Phe           His                                  CH.sub.2                                       CH.sub.2 C(CH.sub.3).sub.2                                                n-Bu       (CH.sub.3).sub.2 CHSO.sub.2                 "             "  "    "        "        ##STR24##                 "             "  "    "        "       (CH.sub.3).sub.2 CHSO.sub.2                 CH.sub.2 CH(CH.sub.2 Ph)CO                               "  "    "        "     ○10       --                  ##STR25##    "  "    "        "       Boc       "             "  "    CH.sub.2 CH.sub.2                                                 ##STR26##       (CH.sub.3).sub.2 CHSO.sub.2                 "             "  "    "        "        ##STR27##                 "             "  "    "        "       (CH.sub.3).sub.2 CHSO.sub.2                 CH.sub.2 CH(CH.sub.2 Ph)CO                               His                                  CH.sub.2                                       CH.sub.2 CH.sub.2                                                 ##STR28##     ○15       --                  ##STR29##    "  "    "        "       Boc       Phe           "  O    "        "       (CH.sub.3).sub.2 CHSO.sub.2                 "             "  "    "        "        ##STR30##                 "             "  "    "        "       (CH.sub.3).sub.2 CHSO.sub.2                 CH.sub.2 CH(CH.sub.2 Ph)CO                               "  "    "        "     ○20       --                  ##STR31##    "  "    "        "       Boc       Phe           His                                  CH.sub.2                                       C(CH.sub.3).sub.2 CH.sub.2                                                 ##STR32##       (CH.sub.3).sub.2 CHSO.sub.2                 "             "  "    "        "        ##STR33##                 "             "  "    "        "       (CH.sub.3).sub.2 CHSO.sub.2                 CH.sub.2 CH(CH.sub.2 Ph)CO                               "  "    "        "     ○25       --                  ##STR34##    "  "    "        "       Boc       Phe           His                                  CH.sub.2                                       CH.sub.2 CH.sub.2                                                 ##STR35##       (CH.sub.3).sub.2 CHSO.sub.2                 "             "  "    "        "        ##STR36##                 "             "  "    "        "       (CH.sub.3).sub.2 CHSO.sub.2                 CH.sub.2 CH(CH.sub.2 Ph)CO                               "  "    "        "     ○30       --                  ##STR37##    "  "    "        "       Boc       Phe           His                                  CH.sub.2                                       CH.sub.2 CH.sub.2                                                 ##STR38##       (CH.sub.3).sub.2 CHSO.sub.2                 "             "  "    "        "        ##STR39##                 "             "  "    "        "       (CH.sub.3).sub.2 CHSO.sub.2                 CH.sub.2 CH(CH.sub.2 Ph)CO                               "  "    "        "     ○35       --                  ##STR40##    "  "    "        ".    __________________________________________________________________________

The pharmaceutically-acceptable salts of the peptides of Formula I (inth form of water- or oil-soluble or dispersible products) include theconventional non-toxic salts or the quarternary ammonium salts of thesepeptides which are formed, e.g., from inorganic or organic acids orbases. Examples of such acid addition salts include acetate, adipate,alginate, aspartate, benzoate, benzenesulfonate, bisulfate, butyrate,citrate, camphorate, camphorsulfonate, c-cylopentan-epropionate,digluconate, dodecylsulfate, ethanesulfonate, fumarate, glucoheptanoate,glycerophosphate, hemisulfate, heptanoate, hexanoate, hydrochloride,hydrobromide, hydroiodide, 2 hydroxyethanesulfonate, lactate, maleate,methanesulfonate, 2 naphthalenesulfonate, nicotinate, oxalate, pamoate,pectinate, persulfate, 3-phenylpropionate, picrate, pivalate,propionate, succinate, tartrate, thiocyanate, tosylate, and undecanoate.Base salts include ammonium salts, alkali metal salts such as sodium andpotassium salts, alkaline earth metal salts such as calcium andmagnesium salts, salts with organic bases such as dicyclohexylaminesalts, N-methyl-D glucamine, and salts with amino acids such asarginine, lysine, and so forth. Also, the basic nitrogen-containinggroups may be quaternized with such agents as lower alkyl halides, suchas methyl, ethyl, propyl, and butyl chloride, bromides and iodides;dialkyl sulfates like dimethyl, diethyl, dibutyl; and diamyl sulfates,long chain halides such as decyl, lauryl, myristyl and stearylchlorides, bromides and iodides, aralkyl halides like benzyl andphenethyl bromides and others.

Chemical synthesis of the compounds with the general structure given informula I may be accomplished in several ways as illustrated by thefollowing generalized procedures (wherein "ACHP", an abbreviation of2-Amino-3-Cyclohexyl 1-HydroxyPropyl, is used in describing thestructural segment which joins the A-B D and ##STR41## portions of theinvention as shown in formula I. The ACHP segment is connected throughthe 1-position of the propyl chain to the 3-position of the piperidinoneor caprolactam, to the 2-position of the morpholinone orthiamorpholinone, and through the amino group to the A-B-D segment.)

Method A:

Step A1. A derivative of 2-piperidinone, caprolactam, 3 morpholinone or3-thiamorpholinone is obtained through commercial sources or is preparedusing well known chemical methods for preparation of members in theseclasses (see Example 1);

Step A2. The enolate of the 2 piperidinone, caprolactam, 3-morpholinoneor 3-thiamorpholinone derivative is generated, such as by using LDA asthe base, and is added to a nitrogen-protected (e.g., N--Boc or N--CBZprotected) α-amino-aldehyde (e.g., N-tert butyloxycarbonylcyclohexylalaninal) to give a Boc- or CBZ-protected amino alcoholderivative (see Example 2B);

Step A3. The nitrogen protecting group of the amino alcohol derivativefrom step A2 is removed (e.g., by hydrogenolysis for CBZ protection, orTFA treatment for Boc protection) and the amine is coupled usingstandard peptide methodology to one or two amino acids, or to anappropriate carboxylic acid, the structure(s) of which is/are describedby A, B, and D in the general formula I (e.g., see Example 3A steps 1and 2); and

Step A4. Removal of any protecting groups which may have been used,e.g., on the lactam substituent(s) R¹³, R¹⁴, or E), or on the amino acidside chains (see Example 3A, step 3), gives the final products.

Method B:

Steps A1 and A2 are followed.

Step B3. Modification of the lactam substituent(s) (R¹³, R¹⁴, and E ingeneric formula I) is/are performed, as shown in Examples 2V-2PP.

Then Steps A3 and A4 are followed.

Method C:

Steps A1, A2 and A3 are followed.

Step C4: Modification of the lactam substituent(s) R¹³, R¹⁴, and E ingeneric formula I) is/are performed, as shown in Examples 3D.

Step A4 is then followed.

The novel peptides of the present invention possess a high degree ofactivity in treating renin-associated hypertension, hyperald-osteronismand/or congestive heart failure in humans, as well as in otherwarm-blooded animals such as mice, rats, horses, dogs and cats.

For these purposes, the peptides of the present invention may beadministered orally, parenterally (including subcutaneous injections,intravenous, intramuscular, intrasternal injection or infusiontechniques), by inhalation spray, or rectally, in dosage unitformulations containing conventional non-toxic, pharmaceuticallyacceptable carriers, adjuvants and vehicles.

Thus, in accordance with the present invention there is further provideda method of treating and a pharmaceutical composition for treatingrenin-associated hypertension, hyperaldosteronism, and/or congestiveheart failure. This treatment involves administering to a patient inneed of such treatment a pharmaceutical composition comprising a-pharmaceutical carrier, optionally with an adjuvant, and atherapeutically-effective amount of a peptide of the formula: ##STR42##wherein A, B, D, R¹, X, Y and E are defined above, or apharmaceutically-acceptable salt thereof.

These pharmaceutical compositions may be in the form oforally-administrable suspensions or tablets; nasal sprays; sterileinjectable preparations, for example, as sterile injectable aqueous oroleagenous suspensions; or suppositories.

When administered orally as a suspension, these compositions may containmicrocrystalline cellulose for imparting bulk, alginic acid or sodiumalginate as a suspending agent, methylcellulose as a viscosity enhancer,and sweetners/flavoring agents known in the art. As immediate releasetablets, these compositions may contain microcrystalline cellulose,dicalcium phosphate, starch, magnesium stearate and lactose and/or otherexcipients, binders, extenders, disintegrants, diluents and lubricantsknown in the art.

When administered by nasal aerosol or inhalation, these compositions maybe prepared as solutions in saline, employing benzyl alcohol or othersuitable preservatives, absorption promoters to enhance bioavailability,flourocarbons, and/or other solubilizinq or dispersing agents known inthe art.

The injectable solutions or suspensions may be formulated according toknown art, using suitable non-toxic, -parenterally-acceptable diluentsor solvents, such as mannitol, 1,3 butanediol, water, Ringer's solutionor isotonic sodium chloride solution, or suitable dispersing or wettingand suspending agents, such as sterile, bland, fixed oils, includingsynthetic mono- or diglycerides, and fatty acids, including oleic acid.

When rectally administered in the form of suppositories, thesecompositions may be prepared by mixing the drug with a suitable nonirritating excipient, such as cocoa butter, synthetic glyceride estersor polyethylene glycols, which are solid at ordinary temperatures, butliquify and/or dissolve in the rectal cavity to release the drug.

Dosage levels of the order of 0.02 to 2.0 grams-per-day are useful inthe treatment of the above-indicated conditions, with oral doses two-tofive times higher. For example, renin-associated hypertension andhyperaldosteronism are effectively treated by the administration of from10 to 50 milligrams of the compound per kilogram of body weight from oneto three times per day. It will be understood, however, that thespecific dose level and frequency of dosage for any particular patientwill depend upon a variety of factors including the activity of thespecific compound employed, the metabolic stability and length of actionof that compound, the age, body weight, general health, sex, diet, modeand time of administration, rate of excretion, drug combination theseverity of the particular condition, and the host undergoing therapy.

The present invention is also directed to combinations of the novelrenin-inhibitory peptides of Formula I with one or more antihypertensiveagents selected from the group consisting of diuretics, α-and/orβ-adrenergic blocking agents, CNS-acting agents, adrenergic neuronblocking agents, vasodilators, angiotensin I converting enzymeinhibitors, calcium channel blockers, and other antihypertensive agents.

For example, the compounds of this invention can be given in combinationwith such compounds or salt or other derivative forms thereof as:

Diuretics: acetazolamide; amiloride; bendro flumethiazide; benzthiazide;bumetanide; chlorothiazide; chlorthalidone; cyclothiazide; ethacrynicacid; furosemide; hydrochlorothiazide; hydro flumethiazide; indacrinone(racemic mixture, or as either the (+) or (-) enantiomer alone, or amanipulated ratio, e.g., 9:1 of said enantiomers, respectively);metolazone; methyclothiazide; muzolimine; polythiazide; guinethazone;sodium ethacrynate; sodium nitroprusside; spironolactone; ticrynafen;triamterene; trichlormethiazide; α-Adrenergic Blocking Agents:dibenamine; phentolamine; -phenoxybenzamine; prazosin; tolazoline;β-Adrenergic Blocking Agents: atenolol; metoprolol; nadolol;propranolol; timolol;

((+)-2-[3-(tert-butylamino) 2-hydroxypropoxy]-2-furananilide)(ancarolol);

(2 acetyl-7-(2-hydroxy-3-isopropy-lamincpropoxy)benzofuran HCl)(befunolol);

((±; 1-(isopropylamino)3-(p-(2-cyclopropylmethoxyethyl)-phenoxy)-2-propranol HCl) (betaxolol);

(1-[(3,4-dimethoxyphenethyl)amino]-3-(m-tolyloxy)-2-propanol HCl)(bevantolol);

((±)-1-(4-((2-isopropoxyethoxy)methyl)phenoxy)-3-isopropylamino-2-propanol)fumarate)(bisoprolol);

(4-(2-hydroxy-3-[4-(phenoxymethyl)-piperidino]-propoxy)-indole);

(carbazolyl-4-oxy-5,2-(2-methoxyphenoxy)-ethylamino-2-propanol);

(1-((1,1-dimethylethyl)amino)-3-((2-methyl-1H-indol-4-yl)oxy)-2-propanolbenzoate) (bopindolol);

(1-(2-exobicyclo[2.2.1]-hept-2-ylphenoxy)3-[(1-methylethyl)amino]-2-propanolHCl) (bornaprolol);

(o[2-hydroxy-3-[(2-indol-3-yl-1,1-dimethylethyl)amino]propoxy]benzonitrileHCl) (bucindolol);

(α[(tert.butylamino)methyl]-7-ethyl-2-benzofuranmethanol) (bufuralol);

(3-[3-acetyl-4-[3-(tert.butylamino)-2-hydroxypropyl]phenyl]1,1-diethylureaHCl) (celiprolol);

((±)-2-[2-[3-[(1,1-dimethylethyl)amino]-2-hydroxypropoxy]phenoxy]-N-methylacetamideHCl) (cetamolol);

(2-benzimidazolyl-phenyl(2-isopropylaminopropanol));

((±) 3'-acetyl-4'-(2-hydroxy-3-isopropylaminopropoxy)acetanilide HCl)(diacetolol);

(methyl-4-[2-hydroxy3-[(1-methylethyl)aminopropoxy]]-benzenepropanoateHCl) (esmolol);

(erythro DL-1-(7-methylindan4-yloxy)-3-isopropylaminobutan 2 ol);

(1-(tert.butylamino)-3-[O-(2-propynyloxy)phenoxy]-2-propanol (pargolol);

(1-(tert.butylamino)-3-[o-(6-hydrazino-3-pyridazinyl)-phenoxy]-2-propanoldiHCl) (prizidilol);

((-)-2-hydroxy-5-[(R)-1-hydroxy-2-[(R)-(1-methyl-3-phenylpropyl)amino]ethyl]benzamide);

(4-hydroxy-9-[2-hydroxy-3-(isopropylamino)-propoxy]-7-methyl-5H-furo[3,2-g][1]benzopyran-5-one)(iprocrolol);

((-)-5-(tert.butylaaino)-2-hydroxypropoxy]-3,4-dihydro1-(2H)-naphthalenoneHCl) (levobunolol);

(4-(2-hydroxy-3-isopropylamino-propoxy)-1,2-benzisothiazole HCl);

(4-[3-(tert.butylamino)-2-hydroxypropoxy]-N-methylisocarbostyril HCl);

((±)-N-2-[4-(2-hydroxy-3-isopropyl aminopropoxy)-phenyl]ethylN'-isopropylurea) (pafenolol);

(3-[[(2-trifluoroacetamido)ethyl]amino]-1-phenoxypropan-2-ol);

(N-(3-(o -chlorophenoxy)-2-hydroxypropyl)-N'-(4'-chloro2,3-dihydro-3-oxo-5-pyridazinyl) ethylenediamine);

((±)N-[3-acetyl-4-[2-hydroxy-3-[(1-methylethyl)amino]-propoxy]phenyl]butanamide)(acebutolol);

((±)-4'-[3-(tert-butylamino)-2-hydroxypropoxy]spiro[cyclohexane1,2'-indan]-1'-one) (spirendolol);

(7-[3-[[2-hydroxy-3-[(2-methylindol-4-yl)oxy]propyl]amino]butyl]thiophylline)(teoprolol);

((±) 1-tert.butylamino-3-(thiochroman-8-yloxy)-2-propanol) (tertatolol);

((±)-1-tert.butylamino-3-(2,3-xylyloxy)-2-propanol HCl) (xibenolol);

(8-[3-(tert.butylamino)-2-hydroxypropoxy]-5-methylcoumarin) (bucumolol);

(2-(3-(tert.butylamino)-2-hydroxy-propoxy)benzonitrile HCl)(bunitrolol);

((±)-2'-[3-(tert-butylamino)-2-hydroxypropoxy-5'-fluorobutyrophenone)(butofilolol);

(1-(carbazol-4-yloxy)-3-(isopropylamino)-2-propanol) (carazolol);

(5-(3-tert.butylamino-2-hydroxy)propoxy-3,4-dihydrocarbostyril HCl)(carteolol);

(1 (tert.butylamino)-3-(2,5-dichlorophenoxy)-2-propanol) (cloranolol);

(1-(inden-4(or 7)-yloxy)-3-(isopropylamino)-2-propanol HCl) (indenolol);

(1-isopropylamino-3-[(2-methylindol-4-yl)oxy]-2-propanol) (mepindolol);

(1-(4-acetoxy-2,3,5-trimethylphenoxy)-3-isopropylaminopropan-2-ol)(metipranolol);

(1-(isopropylamino)-3-(o-methoxyphenoxy)-3-[(1-methylethyl)amino]-2-propanol)(moprolol);

((1-tert.butylamino)-3-[(5,6,7,8-tetrahydro-cis6,7-dihydroxy-1-naphthyl)oxy]-2-propanol) (nadolol)

((S)-1-(2-cyclopentylphenoxy)-3-[(1,1-dimethylethyl)amino]-2-propanolsulfate (2:1)) (penbutolol);

(4'-[1-hydroxy-2-(amino)ethyl]methanesulfonanilide) (sotalol);

(2-methyl-3-[4-(2-hydroxy-3-tert.butylaminopropoxy)-phenyl]-7-methoxy-isoquinolin-1-(2H)-one);

(1-(4-(2-(4-fluorophenyloxy)ethoxy)phenoxy)-3-isopropylamino-2-propanolHCl);

((-)-p-[3-[(3,4-dimethoxyphenethyl)amino]-2-hydroxypropoxy]-β-methylcinnamonitrile)(pacrinolol);

((±)-2-(3'-tert.butylamino-2'-hydroxypropylthio)-4-(5'-carbamoyl-2'-thienyl)thiazole HCl) (arotinolol);

((±)1-[p-[2-(cyclopropylmethoxy)ethoxy]phenoxy]-3-(isopropylamino)-2-propanol)(cicloprolol);

((±)-1-[(3-chloro-2-methylindol-4-yl)oxy]-3-[(2-phenoxyethyl)amino]-2-propanol)(indopanolol);

((±) 6-[[2-[[3-(p-butoxyphenoxy)-2-hydroxypropyl]amino]ethyl]amino]-1,3dimethyluracil) (pirepolcl);

(4-(cyclohexylamino)-1-(1-naphtholenyloxy)-2-butanol);

(1-phenyl-3-2-[3-(2-cyanophenoxy)-2-hydroxypropyl]-aminoethyl]hydantoinHCl);

(3,4-dihydro-8-(2-hydroxy-3-isopropylaminopropoxy)-3-nitroxy-2H-1-benzopyran)(nipradolol);

α- and β-Adrenergic Blocking Agents:

((±)-1-tert-butylamino)-3-[o-[2-(3-methyl5-iso-xazolyl)vinyl]phenoxy]-2-propanol) (isoxaprolol);

(1-isopropylamino-3-(4-(2-nitroxyethoxy)phenoxy)-2-propanol HCl);

(4-hydroxy-α-[[3-(4-methoxyphenyl)-1-methylpropyl]-aminomethyl]-3-(methylsulfinyl)-benzmethanolHCl) (sulfinalol);

(5-[1-hydroxy-2-[[2-(o-methoxyphenoxy)ethyl]amino]-ethyl]-2-methylbenzenesulfonamideHCl);

(5-[1-hydroxy-2-[(1-methyl-3-phenylpropyl)amino]ethyl]-salicylamide HCl)(labetalol);

(1-((3-chloro-2-methyl-1H-indol-4-yl)oxy)-3-((2-phenoxyethyl)amino)-2-propanol-hydrogenmalonate) (ifendolol);

(4-(2-hydroxy-3-[(1-methyl-3-phenylpropyl)amino]-propoxy)benzeneacetamide);

(1-[3-[[3-(1-naphthoxy)-2-hydroxypropyl]-amino]-3,3-dimethyl-propyl]-2-benzimidazolinone);

(3-(1-(2-hydroxy-2-(4-chlorophenylethyl)-4-piperidyl)-3,4-dihydroxy)quinoxolin-2-(1H)-one);

CNS-Acting Agents: clonidine; methyldopa;

Adrenergic Neuron Blocking Agents: quanethidine; reserpine and otherrauwolfia alkaloids such as rescinnamine;

Vasodilators: diazoxide; hydralazine; minoxidil;

Angiotensin I Converting Enzyme Inhibitors:

1-(3-mercapto-2-methyl-1-oxopropyl)-L-proline (captopril);

(1-(4-ethoxycarbonyl-2,4-(R,R)-dimethylbutanoyl)-indoline-2(S)-carboxylicacid);

(2-[2-[[1-(ethoxycarbonyl)-3-phenylpropyl]amino]-1-oxopropyl]-1,2,3,4-tetrahydro-3-isoquinoline carboxylicacid);

((S)-1-[2-[[1-(ethoxycarbonyl)-3-phenylpropyl]amino]-1-oxopropyl]octahydro-1H-indole-2-carboxylicacid HCl);

(Ncyclopentyl-N-(3-(2,2-dimethyl-1-oxopropyl)thiol-2-methyl-1-oxopropyl)glycine)(pivalopril);

((2R,4R)-2-(2-hydroxyphenyl)-3-(3-mercaptopropionyl)-4-thiazolidinecarboxylicacid);

(1-(N[1-(S)-ethoxycarbonyl-3-phenylpropyl]-(S)-alanyl)-cis,syn-octahydroindol-2(S)carboxylic acid HCl);

((-)-(S)-1-[(S)-3-mercapto-2-methyl-1-oxopropyl]-indoline-2-carboxylicacid);

([1(S),4S]-1-[3-(benzoylthio)-2-methyl-1-oxopropyl]-4-phenylthioL-proline;

(3-([1-ethoxycarbonyl-3-phenyl-(1S)-propyl]amino)-2,3,4,5-tetrahydro-2-oxo-1-(3S)-benzazepine-1-acetic acid HCl);

(N-(2-benzyl-3-mercaptopropanoyl)-S-ethyl-L-cysteine) and the S methylanalogue;

(N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanyl-L-proline maleate)(enalapril);

N-[1-(S)-carboxy-3-phenylpropyl]-L-alanyl-1-proline;

N² -[1-(S)-carboxy-3-phenylpropyl]-L-lysyl-L-proline (lysinopril);

Calcium Channel Blockers:α3-[[2-(3,4-dimethoxyphenyl)ethyl]methylamino]propyl]-3,4-dimethoxy-α-(1-methylethyl)benzeneacetonitrile (verapamil);

1,4-dihydro-2,6-dimethyl-4-(2-nitrophenyl)-3,5-pyridinedicarboxylic aciddimethyl ester (nifedipine);

2-(2,2-dicyclohexylethyl)piperidine (perhexiline);

N-(1-methyl-2-phenylethyl)- -phenylbenzenepropanamine (prenylamine);

3-(aminosulfonyl)-4-chloro-N-(2,3-dihydro-2-methyl-1H-indol-1-yl)benzamide(indapamide);

(2'-(2-diethylaminoethoxy)-3-phenylpropiophenone (etafenone);

(4-[4,4-bis-(4-fluorophenyl)butyl]-N-(2,6-dimethylphenyl)-1-piperazineacetamide)(lidoflazine);

(2-(N-benzyl-N-methylamino)ethylmethyl-2,6dimethyl-4-(m-nitrophenyl)-1,4-dihydro-3,5-pyridinedicarboxylate HCl)(nicardipine);

(N-(3,4-dimethoxyphenethyl)-2-(3,4-dimethoxyphenyl)-N-methyl-m-dithiane-2-propylamine-1,1,3,3-tetraoxide)(tiapamil);

(5,6-dimethoxy-2-(3-[(α-(3,4-dimethoxy)phenylethyl)-methylamino]propyl)phthalimidine)(falipamil);

(β-[(2-methylpropoxy)methyl]-N-phenyl-N-phenylmethyl-1-pyrrolidineethanamineHCl monohydrate) (bepridil);

((+)-cis-3-(acetyloxy)-5-[2-(dimethylamino)ethyl]-2,3-dihydro-2-(4-methoxyphenyl)-1,5-benzothiazepin-4-(5H)-one)(diltiazem);

((E)-1-[bis-(p fluorophenyl)methyl]-4-cinnamylpiperazine di HCl)(flunarizine);

(5-[(3,4-dimethoxyphenethyl)methylamino]-2-isopropyl-2-(3,4,5-trimethoxyphenyl)valeronitrile(gallopamil);

(ethylmethyl(2,3-dichlorophenyl)-1,4-dihydro-2,6-dimethyl-3,5-pyridinedicarboxylate(felodipine);

(isopropyl-2-methoxyethyl-1,4-dihydro-2,6-dimethyl-4-(3-nitrophenyl)-3,5-pyridinecarboxylate)(nimodipine);

(3-ethyl-5-methyl-1,4-dihydro-2,6-dimethyl-4-(3-nitrophenyl)-3,5-pyridine-dicarboxylate)(nitrendipine);

Other Antihypertensive Agents: aminophylline; cryptenamine acetates andtannates; deserpidine; meremethoxylline procaine; pargyline;trimethaphan camsylate; and the like, as well as admixtures andcombinations thereof.

Typically, the individual daily dosages for these combinations can rangefrom about one-fifth of the minimally-recommended clinical dosages tothe maximum recommended levels for the entities when they are givenalone. Coadministration is most readily accomplished by combining theactive ingredients into a suitable unit dosage form containing theproper dosages of each. Other methods of coadministration are, ofcourse, possible.

The renin inhibitory novel peptides of the present invention may also beutilized in in vivo or in vitro diagnostic methods for the purpose ofestablishing the significance of renin as a causative or contributoryfactor in hypertension, hyperaldo steronism or congestive heart failurein a particular patient.

In the in vivo method, a novel peptide of the present invention isadministered to a patient, preferably by intravenous injection, althoughparenteral administration is also suitable, at a hypotensive dosagelevel in a single dose of from 0.1 to 10 mg per kg of body weight, andthe resulting transitory fall in blood pressure, if it occurs, indicatessupranormal plasma renin levels.

In vitro methods which may be employed involve incubating a body fluid,preferably plasma, with a novel peptide of the present inventionaccording to methods described in Boger et al., J. Med. Chem., 1985, 28,1779-1790.

The following are intended to exemplify the present invention, without,however, limiting it.

EXAMPLE 1 SYNTHESIS OF STARTING LACTAMS

A. Preparation of 1-Methylcaprolactam

Caprolactam (25 g; 0.22 mol) was added portionwise over a period of 15minutes to a suspension of NaH (10.6 g of a 60% suspension in mineraloil; 0.265 mol) in degassed DMF (500 mL). The mixture was stirred for 3hours at room temperature under an atmosphere of nitrogen and was thencooled to 0° C. Methyl iodide (39.5 g; 0.275 mol) was added dropwise tothe cold suspension over a period of 30 minutes, and stirring wascontinued overnight, allowing the cooling bath to warm to roomtemperature.

Acetic acid (ca. 2 mL) was added to quench any remaining NaH, and thesuspension was filtered, with the bulk of the DMF being removed bydistillation at reducted pressure (ca. 20 torr), until NaI had formed aheavy precipitate. Ether was added and the mixture was cooled forseveral hours in a refrigerator.

The NaI was removed by filtration, and after the ether had been removedunder reduced pressure on the rotovap, distillation under reducedpressure was resumed. 1 Methylcaprolactam (20 g; 71% yield) wascollected, boiling at 110-115° C. (10 torr).

B. Preparation of 4-Methyl-3 Morpholinone

The title compound was prepared from 2 (methylamino)ethanol ndchloroacetyl chloride in basic ethanol using the procedure of Surry, et.al., J. Am. Chem. Soc. (1955) 77, 633. The product was purified byvacuum distillation from calcium hydride, bp 108-112° C. (10 torr).

C. Preparation of 1-Butyl 2-Piperidinone

2-Piperidinone is deprotonated with NaH in DMF solution and butylatedwith n-butyl iodide to give the title compound.

D. Preparation of 1-Butyl-6,6-Dimethyl-2-Piperidinone

The oxime of 2,2 dimethylcyclopentanone is prepared by reaction of theketone with hydroxylamine hydrochloride in EtOH, and treated withp-toluenesulfonyl chloride and DMAP. Heating then effects Beckmannrearrangement to give 6,6-dimethyl-2-piperidinone which is deprotonatedwith NaH in DMF solution. Reaction with n-butyl iodide then gives thetitle compound.

E. Preparation of 1-(2-Diethylaminoethyl)-2-Piperidinon

2-Piperidinone is deprotonated with NaH in DMF solution and reactionwith allyl bromide, and 1-allyl-2-piperidinone is obtained bydistillation. Ozonolysis in MeOH solution at -78° C., followed byreductive workup with Me₂ S, gives the aldehyde derivative, which isreductively aminated using diethylamine hydrochloride and NaBH₄ to givethe title compound.

F. Preparation of 1.5.5 Trimethyl-2-Piperidinone

The oxime of 3,3 dimethylcyclopentanone is prepared by treatment of theketone with hydroxylamine hydrochloride in EtOH. Heating the oxime withp-toluenesulfonyl chloride and DMAP effects Beckmann rearrangement togive a mixture of 4,4-dimethyl 2-piperidinone and the desired5,5-dimethyl-2-piperidinone.

This mixture is separated chromatographically and the5,5-dimethyl-2-piperidinone is methylated by reaction with NaH in DMFsolution followed by treatment with methyl iodide to give the titlecompound.

G. Preparation of 4 Methyl-5-(2-Propyl)-3-Morpholinone

Treatment of valinol in basic ethanol with chloroacetyl chloride usingthe procedure of Surry, et. al., J. Am. Chem. Soc. (1955) 77, 633, gives5-(2-propyl)-3-morpholinone. Deprotonation of the latter with NaH in DMFsolution and reaction with methyl iodide gives the title compound.

H. Preparation of 4-Methyl-3-Thiamorpholinone

Heating N methylaziridine with ethyl mercaptoacetate gives the titlecompound.

EXAMPLE 2 SYNTHESIS OF BOC-(ACHP)-LACTAMS

A. Preparation of Boc-(ACHP)-1-Methyl-2-Piperidinone using a TypicalLactam Aldol Addition Procedure

To a 0° C. solution of diisopropylamine (4.11 g; 40.7 mmol) in dry THF(120 mL) under an atmosphere of nitrogen was added n-butyllithium (25.1mL of a 1.6 M solution in hexane: 40.1 mmol). After being stirred for 10minutes, the resulting solution was cooled to -78° C., at which time asolution of 1-methyl2-piperidinone (4.52 g, 40.0 mmol) in dry THF (10mL) was added dropwise over a period of 5 minutes.

The resulting solution was stirred at -78° C. for 1.5 hour, when a 78°C. solution of N-Boc-L-cyclohexylalaninal (10.2 g; 40.0 mmol), preparedaccording to the method of Boger, et. al., J. Med. Chem., (1985) 28,1779, in dry THF (60 mL) was added rapidly via cannula. After beingstirred for 5 minutes at -78° C., the reaction was quenched by theaddition of 10 mL of water, and the cooling bath was removed and morewater (25 mL) and ether (200 mL) were added.

The mixture was extracted with 5% aqueous HCl (200 mL), saturated withaqueous NaHCO₃ (200 mL), and dried (MgSO₄) and filtered. Removal of thesolvents under reduced pressure gave a viscous oil, from which thediasteriomeric aldol products was separated by flash chromatography(SiO₂ ; 2%-5% MeOH/CH₂ Cl₂).

The 2S,3S,4S diasteriomer was obtained as a viscous oil (2.80 g; 19%yield).

B. Preparation of Boc (ACHP)-4-Methyl-3-Morpholinone

The title compound (2S,3S,4S diasteriomer) was obtained by aldolreaction of Boc-L-cyclohexylalaninal and 4 methyl 3 morpholinone usingthe procedure described in Example 2A.

C. Preparation of Boc-(ACHP)-1-Methylcaprolactam

The title compound (2S,3S,4S, diasteriomer) was obtained by aldolreaction of Boc-L cyclohexylalaninal and 1-methylcaprolactam using theprocedure described in Example 2A.

The following Boc--(ACHP)--lactams are prepared using the aldolprocedure described in Example 2A

D. Boc-(ACHP 1-Butyl-2-Piperidinone

E. Boc-(ACHP)-1-(2-Diethylaminoethyl)-2-Piperidinone

F. Boc-(ACHP) 1-Butyl-6,6-Dimethyl-2-Piperidinone

G. Boc-(ACHP)-1,5,5 Trimethyl-2-piperidinone

H. Boc-(ACHP)-4-Methyl-3-thiamorpholinone

I. Boc-(ACHP)-4-Methyl-5-(2-Propyl)-3-Morpholinone

EXAMPLE 3 AMINO ACID COUPLING PROCEDURES AND SUBSEQUENT TRANSFORMATIONSA. Preparation of Boc-Phe-His-(ACHP)-1-Methyl-2-Piperidinone Step 1.Boc-(DNP)His-(ACHP)-1-Methyl-2-Piperidinone

To a solution of Boc (ACHP) 1 methyl 2-piperidinone (2.20 g; 5.98 mmol)in CH₂ Cl₂ (4 mL) was added TFA (4 mL) and the solution was stored underan atmosphere of nitrogen at ambient temperature for 45 minutes. ExcessTFA and solvent were removed under reduced pressure and the oil soobtained was triturated in ether, giving a white solid which wascollected by filtration, washed with ether, and dried under reducedpressure.

The TFA salt (2.10 g; 92% yield) was dissolved under an atmosphere ofnitrogen in a minimum amount (5-6 mL) of DMF (more ideally in EtOAc ifthe salt had been soluble) and stored while the activation ofBoc(DNP)His--OH was accomplished by in situ formation of a mixedanhydride, as described immediately below.

(Activation of Boc(DNP)His--OH) To a suspension of Boc(DNP)His OH (3.23g; 7.69 mmol) in dry EtOAc (30 mL) under an atmosphere of nitrogen wasadded NMM (930 ul; 8.45 mmol), which caused dissolution of any remainingsolid. The solution was cooled to -23° C. at which point isobutylchloroformte (957 ul; 7.38 mmol) was added, and the resulting solutionwas stirred at -23° C. for 25 minutes.

After completion of the activation, the DMF solution of the TFA salt wasneutralized by the addition of NMM (812 ul; 7.38 mmol) and was added viacannula to the cold solution of the mixed anhydride. After being stirredfor 1 hour at -23° C., the reaction mixture was warmed to 0° C. and wasstirred for 1.5 hour. The cooling bath was removed and stirring of themixture was continued for another 3 hours, at which time the reactionwas quenched by the addition of water (50 mL).

The mixture was diluted with EtOAc (125 mL) and was washed successivelywith 5% aqueous HCl (150 mL), water (20 mL), and saturated aqueousNaHCO₃ (200 mL). The organic layer was dried (MgSO₄), filtered, andconcentrated under reduced pressure to give an orange solid which wasflash chromatographed (SiO₂ ; 3%-5% MeOH/CH₂ Cl₂).

The coupling product was obtained as a yellow solid (3.05 g; 75% yield).

Step 2 Boc-Phe-(DNP)His (ACHP)-1-Methyl-2-Piperidinone

To a solution of the coupling product obtained from Step 1 (3.05 g; 4.55mmol) in CH₂ Cl₂ (4 mL) was added TFA (4 mL), and the mixture was storedunder an atmosphere of nitrogen for 45 minutes, at which time thesolvent and excess TFA were removed under reduced pressure. The yellowoil so obtained was triturated in ether and the solid which resulted wascollected by filtration, washed with ether, and dried under reducedpressure.

The resulting orange yellow solid (2.99 g; 96% yield) was dissolved,under an atmosphere of nitrogen, in dry EtOAc (20 mL) and stored whilethe activation of Boc-Phe-Oh was accomplished by in situ formation of amixed anhydride as described below.

(Activation of Boc--Phe--OH) To a suspension of Boc--Phe--OH (1.52 g;5.74 mmol) in dry EtOAc (25 mL) under an atmosphere of nitrogen wasadded NMM (695 μl; 6.31 mmol). The resulting solution was cooled to -23°C., isobutyl chloroformate (715 ul; 5.51 mmol) was added, and thesolution was stirred for 25 minutes.

After completion of the activation, the solution of the TFA salt wasneutralized by the addition of NMM (555 μl; 5.06 mmol) and was added viacannula to the cold solution of the mixed anhydride. The resultingmixture was stirred at -23° C. for 1 hour, at 0° C. for 1.5 hour, andthen at room temperature for 3.5 hours, and the reaction was quenched bythe addition of water (50 mL), and was diluted with EtOAc (125 mL).

The organic phase was washed successively with 5% aqueous HCl (150 mL),water (20 mL) and saturated aqueous NaHCO₃ (150 mL), then dried (MgSO₄),and filtered. Removal of the solvent under reduced pressure gave a solidwhich was flash-chromatographed (SiO₂ ; 4%-7% MeOH/CH₂ Cl₂). Thecoupling product was obtained as a yellow solid (3.24 g; 87% yield).

Step 3. Boc-Phe-His-(ACHP)-1-Methyl-2-Piperidinone

The coupling product obtained from Step 2 (3.24 g; 3.96 mmol) wasdissolved under an atmosphere of nitrogen in dry CH₂ Cl₂ (8 mL), thenthiophenol (4 mL) and NMM (60 μl; 0.54 mmol) were added. The mixture wasstirred at room temperature for 5 hours, at which time the solvent andexcess thiophenol were removed under reduced pressure (0.2 torr) at 30°C. The residue was flash chromatoqraphed (SiO₂ ; 5%-10% MeOH/CH₂ Cl₂),giving the title compound as an off-white powder (2.27 g; 88% yield).

The product exhibited satisfactory NMR spectral properties, was analzyedfor purity by reverse phase HPLC, and was anlyzed for composition usingC, H, and N combustion analysis. Inhibition of human plasma renin wasassayed using the in vitro method described by Boger, et. al., J. Med.Chem. (1985) 28, 1779. IC₅₀ =47 nM.

B. Preparation of Boc-Phe His-(ACHP)-4-Methyl-3-Morpholinone

Amino acid coupling was accomplished as described in Example 3A. IC₅₀=31 nM.

C. Preparation of Boc Phe His-(ACHP) 1-Methylcaprolactam

Amino acid coupling was accomplished as described in Example 3A. IC₅₀=203 nM.

D. Preparation ofBoc-Phe-His-(ACHP)-1-(N-Benzyl-2-Diethylaminoethyl)-2-PiperidinoneAcetate

Boc-Phe-His-(ACHP)-1-(Diethylaminoethyl)-2-piperidinone is protected onthe histidine side chain by reaction with (BOC)₂ O in DMF solution. Anexcess of benzyl bromide is added and the mixture is warmed to effectquaternization of the diethylaminoethyl substituent. The Boc group onthe histidine side chain is removed in situ by the addition of water andtriethylamine.

The crude product is purified by preparative reverse phase HPLC and ispassed through an ion exchange column (Bio Rad AG3-X4A resin, acetateform) to produce the title compound. Ipoc--Phe--His--peptides are madeusing sequential coupling, using Ipoc--Phe instead of Boc--Phe, in themethod according to Example 3A, or by fragment coupling, usingIpoc--Phe--His in place of Boc--Phe--His, in the method according toExample 3B. ##STR43## peptides are made by sequential coupling usingmixed anhydride coupling (according to Example 3A) for the histidinegroup, and using mixed anhydride or DCC/HOBT coupling for the ##STR44##group. (CH₃)₂ CH--SO₂ --Phe--His--peptides are made by sequentialcoupling using mixed anhydride coupling (according to Example 3A) forthe histidine group, and using mixed anhydride or DCC/HOBT coupling forthe (CH₃)₂ CH-SO₂ -Phe group.

(CH₃)₂ CH--SO₂ --CH₂ -CH(CH₂ Ph)--CO--His--peptides are made bysequential coupling using mixed anhydride coupling (according to Example3A) for the histidine group and DCC/HOBT or DCC/HOSU coupling for the(CH₃)₂ CH--SO₂ --CH₂ -CH(CH₂ Ph)--CO--group.

2-Indolyl-CO His peptides are made by sequential coupling using mixedanhydride coupling (according to Example 3A) for the histidine group,and DCC or EDC coupling of indole-2-carboxylic acid.

These methods are applied to any of the 9 Boc-(ACHP)-lactams describedin Example 2.

Claims to the Invention follow.

What is claimed is:
 1. A peptide of the formula: ##STR45## wherein: A ishydrogen; C₁ -C₆ -alkyl; aryl, where aryl is unsubstituted or mono-, di-or trisubstituted phenyl, wherein the substituent(s) is/areindependently selected from the group consisting of C₁ -C₇ alkyl, amino,mono- or di-C₁ -C₄ -alkylamino, amino-C₁ -C₄ -alkyl, hydroxy-C₁ -C₄-alkyl, phenyl-C₁ -C₄ -alkyl, mono- or di-C₁ -C₄ -alkylamino-C₁ -C₄-alkyl, guanidyl, guanidyl-C₁ -C₄ -alkyl, hydroxyl, C₁ -C₄ -alkoxy,trifluoromethyl, halo, CHO, --CO₂ H, --CONH₂, --CONH-C₁ -C₄ -alkyl,--CON(C₁ -C₄ -alkyl)₂, --CO--C₁ -C₄ -alkyl, --(CH₂)_(m) -.sup.⊕ N(R³)₂R⁴ A.sup.⊖, where R³ is C₁ -C₄ -alkyl, --(CH₂)₄ --, --(CH₂)₅ - or--(CH₂)₂ -O-(CH₂)₂ -; R⁴ is C₁ -C₄ -alkyl, C₁ -C₄ -hydroxyalkyl, C₁-C.sub. 4 -carboxyalkyl, or --CH₂ -phenyl; A.sup.⊖ is a counterionselected from the group consisting of single negatively charged ions;and m is 0-to-3; --CO₂ --C₁ -C₄ -alkyl, --CO₂ --C₁ -C₄ -alkoxy-C₂ -C₄-alky ##STR46## where A⁻ and m are as defined above, and --NR⁵ R⁶, whereR⁵ and R⁶ are independently hydrogen, unsubstituted or monosubstitutedC₁ -C₄ -alkyl, wherein the substituent is amino, mono- or di-C₁ -C₄ -alkylamino or --.sup.⊕ N(R³)₂ R⁴ A.sup.⊖, where R³, R⁴ and A.sup.⊖ areas defined above; Het, where Het is an unsubstituted or mono-ordisubstituted 5-or 6-membered mono- or bicyclic or benzofused 5-or6-membered heterocyclic ring, where the one or two heteroatoms areindependently selected from the group consisting of N, O, S, NO, SO, SO₂or quaternized N, and the substituent(s) is/are independently selectedfrom the group consisting of hydroxyl, thiol, C₁ -C₆ -alkyl, CF₃, C₁ -C₄-alkoxy, halo, aryl, as defined above, aryl-C₁ -C₄ -alkyl, amino, mono-or diC₁ -C₄ -alkylamino, amino-C₁ -C₄ -alkyl, hydroxy-C₁ -C₄ -alkyl,mono or di-C.sub. 1 -C₄ -alkylamino-C₁ -C₄ -alkyl, guanidyl, guanidylC₁-C₄ -alkyl CHO, CO₂ H, CO₂ -C₁ -C₄ -alkyl, CONH₂, COHN-C₁ -C₄ -alkyl,CON(C₁ -C₄ -alkyl)₂, NR⁵ R⁶, ##STR47## A.sup.⊕ and --(CH₂)_(m) --.sup.⊕N(R³)₂ (R⁴ A.sup.⊖, wherein R⁵, R⁶, A.sup.⊖, m, R³ and R⁴ are as definedabove, or when the heteroatom is N, the substituents are alternatively--(CH₂)q--or --(CH₂)₂ -O-(CH₂)₂ -13 and form a ring with the N-atom,wherein q is 3-to-6; ##STR48## where R² is C₁ -C₇ -alkyl; hydrogen; Het,as defined above; aryl, as defined above; mono-substituted C₁ -C₅-alkyl, wherein the substituent is selected from the group consisting ofaryl, as defined above; Het, as defined above; hydroxyl; CO₂ H; CO₂R⁷,where R⁷ is C₁ -C₅ -alkyl, aryl, as defined above, and aryl-C₁ -C₄-alkyl; CONH₂ ; --CONH-R⁷ or --S(O)_(n) --R⁷, wherein n is 0-to-2 and R⁷is as defined above; C₁ -C₄ -alkoxy; C₃ -C₇ -cycloalkyl; amino; mono- ordi-C₁ -C₄ -alkylamino; NH-aryl, --NH--CH₂ -aryl or --CO--aryl, wherearyl is as defined above; and --NH-Het, --NH--CH₂ --Het or --CO--Het,where Het is as defined above; ##STR49## where R⁷ is as defined above;or ##STR50## where R⁹ is C₁ -C₅ -alkyl, aryl, as defined above, or Het,as defined above; B and D are independently ##STR51## where R¹² ishydrogen, C₁ -C₅ -alkyl or CH₂ -aryl, wherein aryl is as defined above;and R² is as defined above; ##STR52## where R² is as defined above; oreither B or D, but not both simultaneously, is absent;R¹ is hydrogen; C₃-C₆ -alkyl; aryl, as defined above; unsubstituted, mono-, di- ortrisubstituted C₃ -C₇ -cycloalkyl, where the substituent(s) is/areselected from the group consisting of C₁ -C₄ -alkyl, trifluoromethyl,hydroxyl, C₁ -C₄ -alkoxy and halo; or unsubstituted or 4 monosubstituted1,3-dithiolan-2-yl or unsubstituted or 4-mono-substituted1,3-dithian-2-yl, where the substituent is --(CH₂)_(m) -aryl, where mand aryl are as defined above; ##STR53## where R¹³ and R¹⁴ areindependently hydrogen; C₁ -C₇ -alkyl; C₂ -C₇ -alkenyl; --CO₂ H; --CONH₂; --CO₂ R⁷, --CO--NH--R⁷ or --CO--N(R⁷)₂, wherein R⁷ is as definedabove; mono-substituted C₁ -C₅ -alkyl, wherein the substituent isselected from the group consisting of azido; halo; hydroxy; C₁ -C₅-alkoxy; aryl, aryl--CH₂ O, aryloxy, aryl--COO--, aryl--CH₂ --NH--orarylamino, where aryl is as defined above; C₁ -C₅ -alkyl--CO₂ --; R⁷NH--COO--, R⁷ --CO--NH--, R⁷ --NH--CO--NH--or R⁷ --S(O)_(n), where n andR⁷ are as defined above; amino; mono- or di-C₁ -C₄ -alkylamino; or Het,as defined above; or R¹³ and R¹⁴ are connected to form a polymethylenechain of the formula, --(CH₂)_(p), where p is 2-to-6; or ##STR54## whereR¹³ and R¹⁴ are as defined above; Y is CH₂, O, S, SO or SO₂ ; or Y--X is--(CH₂)₄ --; and E is hydrogen; aryl, as defined above; Het, as definedabove; C₂ -C₇ -alkenyl; or unsubstituted or mono- substituted C₁ -C₇-alkyl or unsubstituted or mono-substituted C₃ -C₇ -cycloalkyl, wherethe substituent is selected from the group consisting of aryl,--CO-aryl, --NH-aryl or --O-aryl, wherein aryl is as defined above; Het,--NH-Het, --O-Het, --CO-Het, --NH--CO-Het, CO--NH-Het, CO--NH--CH₂ -Hetor O--CO-Het, wherein Het is as defined above; azido; C₃ -C₇-cycloalkyl; halo; hydroxyl; C₁ -C₄ -alkoxy; --COOH; --O-CO-R⁷,--O-CO-NH-R⁷, --NH-CO-R⁷, --NH-CO-NH-R⁷, --S(O)_(n) --R⁷, --CO₂ R⁷ or--CO-NH-R⁷, wherein R⁷ and n are as defined above; amino; mono- or di-C₁-C₄ -alkylamino; -CHO; and --.sup.⊕ N(R³)₂ R⁸ A.sup.⊖, ##STR55## whereC₁ -C₄ -alkyl, C₁ -C₄ -hydroxyalkyl, C₁ -C₄ -carboxyalkyl, --CH₂ -aryl,wherein aryl is as defined above, or --CH₂ -Het, wherein Het is asdefined above, and R³ and A.sup.⊖ are as defined above;or apharmaceutically-acceptable salt thereof.
 2. A peptide according toclaim 1, wherein A is R² --CO--, R⁹ --SO₂ --, C₁ -C₄ -alkyl-O-CO- or R⁷--NH-CO-, wherein R², R⁷ and R⁹ are as defined in claim 1; B is absentwhen D is L-histidyl or L-valinyl, or B is unsubstituted ormonosubstituted L-phenylalanyl, wherein the substituent is on the phenylring and is para-methoxy or ##STR56## D is absent, when B isunsubstituted or monosubstituted L-phenylalanyl, or D is L-histidyl orL-valinyl; R¹ is cyclohexyl; Y is O or CH₂ ; R¹³ and R¹⁴ in eitherdefinition of X are simultaneously or independently hydrogen or methyl;and E is C₁ -C₆ -alkyl, --(CH₂)_(r) --.sup.⊕ N(R³)₂ R⁸ CH₃ CO.sup.⊖₂,wherein r is 2 or 3 and R³ and R⁸ are as defined in claim 1, ##STR57##wherein R⁸ is as defined in claim
 1. 3. A peptide according to claim 1,having the structure: ##STR58## wherein in the structures:

    __________________________________________________________________________    A            B             D  Y    X        E    __________________________________________________________________________     ○1       Boc       Phe           His                                  CH.sub.2                                       CH.sub.2 CH.sub.2                                                n-Bu       (CH.sub.3).sub.2 CHSO.sub.2                 "             "  "    "        "        ##STR59##                 "             "  "    "        "       (CH.sub.3).sub.2 CHSO.sub.2                 CH.sub.2 CH(CH.sub.2 Ph)CO                               "  "    "        "     ○5       --                  ##STR60##    "  "    "        "       Boc       Phe           His                                  CH.sub.2                                       CH.sub.2 C(CH.sub.3).sub.2                                                n-Bu       (CH.sub.3).sub.2 CHSO.sub.2                 "             "  "    "        "        ##STR61##                 "             "  "    "        "       (CH.sub.3).sub.2 CHSO.sub.2                 CH.sub.2 CH(CH.sub.2 Ph)CO                               "  "    "        "     ○10       --                  ##STR62##    "  "    "        "       Boc       "             "  "    CH.sub.2 CH.sub.2                                                 ##STR63##       (CH.sub.3).sub.2 CHSO.sub.2                 "             "  "    "        "        ##STR64##                 "             "  "    "        "       (CH.sub.3).sub.2 CHSO.sub.2                 CH.sub.2 CH(CH.sub.2 Ph)CO                               His                                  CH.sub.2                                       CH.sub.2 CH.sub.2                                                 ##STR65##     ○15       --                  ##STR66##    "  "    "        "       Boc       Phe           "  O    "        "       (CH.sub.3).sub.2 CHSO.sub.2                 "             "  "    "        "        ##STR67##                 "             "  "    "        "       (CH.sub.3).sub.2 CHSO.sub.2                 CH.sub.2 CH(CH.sub.2 Ph)CO                               "  "    "        "     ○20       --                  ##STR68##    "  "    "        "       Boc       Phe           His                                  CH.sub.2                                       C(CH.sub.3).sub.2 CH.sub.2                                                 ##STR69##       (CH.sub.3).sub.2 CHSO.sub.2                 "             "  "    "        "        ##STR70##                 "             "  "    "        "       (CH.sub.3).sub.2 CHSO.sub.2                 CH.sub.2 CH(CH.sub.2 Ph)CO                               "  "    "        "     ○25       --                  ##STR71##    "  "    "        "       Boc       Phe           His                                  CH.sub.2                                       CH.sub.2 CH.sub.2                                                 ##STR72##       (CH.sub.3).sub.2 CHSO.sub.2                 "             "  "    "        "        ##STR73##                 "             "  "    "        "       (CH.sub.3).sub.2 CHSO.sub.2                 CH.sub.2 CH(CH.sub.2 Ph)CO                               "  "    "        "     ○30       --                  ##STR74##    "  "    "        "       Boc       Phe           His                                  CH.sub.2                                       CH.sub.2 CH.sub.2                                                 ##STR75##       (CH.sub.3).sub.2 CHSO.sub.2                 "             "  "    "        "        ##STR76##                 "             "  "    "        "       (CH.sub.3).sub.2 CHSO.sub.2                 CH.sub.2 CH(CH.sub.2 Ph)CO                               "  "    "        "     ○35       --                  ##STR77##    "  "    "        ".    __________________________________________________________________________


4. A pharmaceutical composition for renin-associated hypertension orcongestive heart failure comprising a pharmaceutical carrier and atherapeutically-effective amount of a peptide according to claim
 1. 5. Apharmaceutical composition according to claim 4, also comprising anadjuvant.
 6. A pharmaceutical composition according to claim 4, alsocomprising one or more compounds selected from the group consistingof:Diuretics: acetazolamide; amiloride; bendroflumethiazide;benzthiazide; bumetanide; chlorothiazide; chlorthalidone; cyclothiazide;ethacrynic acid; furosemide; hydrochlorothiazide; hydroflumethiazide;indacrinone (racemic mixture, or as either the (+) or (-) enantiomeralone, or a manipulated ratio, e.g., 9:1 of said enantiomers,respectively); metolazone; methyclothiazide; muzolimine; polythiazide;quinethazone; sodium ethacrynate; sodium nitroprusside; spironolactone;ticrynafen; triamterene; trichlormethiazide; α-Adrenergic BlockingAgents: dibenamine; phentolamine; phenoxybenzamine; prazosin;tolazoline; β-Adrenergic Blocking Agents: atenolol; metoprolol; nadolol;propranolol; timolol;((±)-2-[3-(tertbutylamino)-2-hydroxypropoxy]-2-furananilide) (ancarolol);(2-acetyl-7-(2-hydroxy-3-isopropylaminopropoxy)benzofuran HCl)(befunolol); ((±)-1-(isopropylamino)-3-(p(2-cyclopropylmethoxyethyl)-phenoxy)-2-propranol HCl) (betaxolol);(1-[(3,4-dimethoxyphenethyl)amino]-3-(m-tolyloxy)-2-propanol HCl)(bevantolol);((±)-1-(4-((2-isopropoxyethoxy)methyl)phenoxy)-3-isopropylamino-2-propanol)fumarate)(bisoprolol);(4-(2-hydroxy-3-[4-(phenoxymethyl)-piperidino]-propoxy)-indole);(carbazolyl-4-oxy-5,2-(2-methoxyphenoxy)-ethylamino-2-propanol);(1-((1,1-dimethylethyl)amino)-3-((2-methyl-1H-indol-4-yl)oxy)-2-propanolbenzoate) (bopindolol);(1-(2-exobicyclo[2.2.1]-hept-2-ylphenoxy-)-3-[(1-methylethyl)-amino]-2-propanolHCl) (bornaprolol);o-[-hydroxy-3-[(2-indol-3-yl-1,1-dimethylethyl)-amino]propoxy]benzonitrileHCl) (bucindolol);(α-[(tert.butylamino)methyl]-7-ethyl-2-benzofuranmethanol) (bufuralol);(3-[3-acetyl-4-[3-(tert.butylamino)-2-hydroxypropyl]-phenyl]-1,1-diethylureaHCl) (celiprolol);((±)-2-[2-[3-[(1,1-dimethylethyl)amino]-2-hydroxypropoxy]phenoxy]N-methylacetamideHCl) (cetamolol); (2-benzimidazolyl phenyl(2-isopropylaminopropanol));((±) 3'-acetyl-4'-(2-hydroxy-3-isopropylaminopropoxy)-acetanilide HCl)(diacetolol);(methyl-4-[2-hydroxy-3-[(1-methylethyl)aminopropoxy]]-benzenepropanoateHCl) (esmolol);(erythro-DL-1-(7-methylindan-4-yloxy)-3-isopropylaminobutan-2-ol);(1-(tert.butylamino)-3-[O-(2-propynyloxy)phenoxy]-2-propanol (pargolol);(1-(tert.butylamino)-3-[o-(6-hydrazino-3-pyridazinyl)-phenoxy]-2-propanoldiHCl) (prizidilol);((-)-2-hydroxy-5-[(R)-1-hydroxy-2-[(R)-(1-methyl-3-phenylpropyl)amino]ethyl]benzamide);(4-hydroxy-9-[2-hydroxy-3-(isopropylamino)-propoxy]-7-methyl-5H-furo[3,2-g][1]-benzopyran-5-one)(iprocrolol);((-)-5-(tert.butylamino)-2-hydroxypropoxy]-3,4-dihydro-1-(2H)-naphthalenoneHCl) (levobunolol);(4-(2-hydroxy-3-isopropylamino-propoxy)-1,2-benzisothiazole HCl);(4-[3-(tert.butylamino)-2-hydroxypropoxy]-N-methylisocarbostyril HCl);((±)-N-2-[4-(2-hydroxy-3-isopropylaminopropoxy)-phenyl]ethyl-N'-isopropylurea) (pafenolol);(3-[[(2-trifluoroacetamido)ethyl]amino]-1-phenoxypropan2-ol);(N-(3-(o-chlorophenoxy)-2-hydroxypropyl)-N'-(4'-chloro2,3-dihydro-3-oxo-5-pyridazinyl)ethylenediamine);((±)-N-[3-acetyl-4-[2-hydroxy-3-[(1-methylethyl)amino]propoxy]phenyl]butanamide)(acebutolol); ((±)-4'-[3-(tertbutylamino)-2-hydroxypropoxy]spiro-cyclohexane-1,2'-indan]-1'-one)(spirendolol);(7-[3-[[-2-hydroxy-3-[(2-methylindol-4-yl)oxy]propyl]-amino]butyl]thiophylline)(teoprolol); ((±)-1-tert.butylamino-3-(thiochroman-8-yloxy)-2-propanol)(tertatolol); ((±) 1-tert.butylamino-3-(2,3-xylyloxy)-2-propanol HCl)(xibenolol); (8-[3-(tert.butylamino)-2-hydroxypropoxy]-5-methylcoumarin)(bucumolol); (2-(3-(tert.butylamino)-2-hydroxy-propoxy)benzonitrile HCl)(bunitrolol);((±)-2'-[3-(tert-butylamino)-2-hydroxypropoxy-5'-fluorobutyrophenone)(butofilolol); (1-(carbazol-4-yloxy)-3-(isopropylamino)-2-propanol)(carazolol);(5-(3-tert.butylamino-2-hydroxy)propoxy-3,4-dihydrocarbostyril HCl)(carteolol); (1-(tert.butylamino)-3-(2,5-dichlorophenoxy)-2-propanol)(cloranolol); (1-(inden-4-(or 7)-yloxy)-3-(isopropylamino)-2-propanolHCl) (indenolol);(1-isopropylamino-3-[(2-methylindol-4-yl)oxy]-2-propanol) (mepindolol);(1-(4-acetoxy-2,3,5-trimethylphenoxy)-3-isopropylaminopropan-2-ol)(metipranolol);(1-(isopropylamino)-3-(o-methoxyphenoxy)-3-[(1-methylethyl)amino]-2-propanol)(moprolol);((1-tert.butylamino)-3-[(5,6,7,8-tetrahydro-cis-6,7-dihydroxy-1-naphthyl)oxy]-2-propanol)(nadolol)((S)-1-(2-cyclopentylphenoxy)-3-[(1,1-dimethylethyl)amino]-2-propanolsulfate (2:1)) (penbutolol);(4'-[1-hydroxy-2-(amino)ethyl]methanesulfonanilide) (sotalol):(2-methyl-3-[4-(2-hydroxy-3-tert.butylaminopropoxy)phenyl]-7-methoxy-isoquinolin-1-(2H)-one);(1-(4-(2-(4-fluorophenyloxy)ethoxy)phenoxy)-3-isopropylamino-2-propanolHCl);((-)-p-[3-[(3,4-dimethoxyphenethyl)amino]-2-hydroxypropoxy]-β-methylcinnamonitrile)(pacrinolol);((±)-2-(3'-tert.butylamino-2'-hydroxypropylthio)4-(5'-carbamoyl2'-thienyl)thiazoleHCl) (arotinolol);((±)-1-[p-[2-(cyclopropylmethoxy)ethoxy]phenoxy]-3-(isopropylamino)-2-propanol)(cicloprolol);((±)-1-[(3-chloro-2-methylindol-4-yl)oxy]-3-[(2-phenoxyethyl)amino]-2-propanol)(indopanolol);((±)-6-[[2-[[3-(p-butoxyphenoxy)-2-hydroxypropyl]-amino]ethyl]amino]-1,3-dimethyluracil)(pirepolol); (4-(cyclohexylamino)-1-(1-naphtholenyloxy)-2-butanol);(1-phenyl-3-[2-[3-(2-cyanophenoxy)-2-hydroxypropyl]aminoethyl]hydantoinHCl);(3,4-dihydro-8-(2-hydroxy-3-isopropylaminopropoxy)-3-nitroxy-2H-1-benzopyran)(nipradolol); α- and β-Adrenergic BlockingAgents:((±)-1-tert-butylamino)-3-[o-[2-(3-methyl-5-isoxazolyl)vinyl]phenoxy]-2-propanol)(isoxaprolol);(1-isopropylamino-3-(4-(2-nitroxyethoxy)phenoxy)-2-propanol HCl);(4-hydroxy-α-[[3-(4-methoxyphenyl)-1-methylpropyl]aminomethyl]-3-(methylsulfinyl)-benzmethanol(HCl) (sulfinalol);(5-[1-hydroxy-2-[[2-(o-methoxyphenoxy)ethyl]amino]ethyl]-2-methylbenzenesulfonamideHCl); (5-[1-hydroxy-2-[(1-methyl-3-phenylpropyl)amino]ethyl]salicylamideHCl) (labetalol);(1-((3-chloro-2-methyl-1H-indol-4-yl)oxy)-3-((2-phenoxyethyl)amino)-2-propanolhydrogenmalonate) (ifendolol);(4-(2-hydroxy-3-[(1-methyl-3-phenylpropyl)amino]-propoxy)benzeneacetamide);(1-[3-[[3-(1-naphthoxy-)-2-hydroxypropyl]-amino]-3,3-dimethyl-propyl]-2-benzimidazol(3-(1-(2-hydroxy-2-(4-chlorophenylethyl)-4-piperidyl)-3,4-dihydroxy)quinoxolin-2(1H)-one); CNS-Acting Agents: clonidine; methyldopa; Adrenergic NeuronBlocking Agents: quanethidine; reserpine and other rauwolfia alkaloidssuch as rescinnamine; Vasodilators: diazoxide; hydralazine; minoxidil;Angiotensin I Converting EnzymeInhibitors:1-(3-mercapto-2-methyl-1-oxopropyl)-L-proline (captopril);(1-(4-ethoxycarbonyl-2,4-(R,R)-dimethylbutanoyl)-indoline-2(S)-carboxylicacid);(2-[2-[[1-(ethoxycarbonyl)-3-phenyl-propyl]amino]-1-oxopropyl]-1,2,3,4-tetrahydro-3-3-isoquinolinecarboxylic acid);((S)-1-[2-[[1-(ethoxycarbonyl)-3-phenylpropyl]amino]-1-oxopropyl]octahydro-1H-indole-2-carboxylic acid HCl);(N-cyclopentyl-N-(3-(2,2-dimethyl-1-oxopropyl)thiol-2-methyl-1-oxopropyl)glycine)(pivalopril);((2R,4R)-2-(2-hydroxyphenyl)-3-(3-mercaptopropionyl)-4-thiazolidinecarboxylicacid);(1-(N-[1(S)-ethoxycarbonyl-3-phenylpropyl]-(S)-alanyl)-cis,syn-octahydroindol-2(S)-carboxylicacid HCl); ((-)(S)-1-[(S)-3-mercapto-2-methyl-1-oxopropyl]indoline-2-carboxylic acid);([1(S),4S]-1-[3-(benzoylthio)-2-methyl-1-oxopropyl]-4-phenylthio-L-proline;(3-([1-ethoxycarbonyl-3-phenyl-(1S)propyl]amino)-2,3,4,5-tetrahydro-2-oxo-1-(3S) -benzazepine-1-acetic acidHCl); (N-(2-benzyl-3-mercaptopropanoyl)-S-ethyl L-cysteine) and the Smethyl analogue;(N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanyl-L-proline maleate)(enalapril); N-[1-(S)-carboxy-3-phenylpropyl]-L-alanyl-1-proline; N²-[1-(S)-carboxy-3-phenylpropyl]-L-lysyl-L-proline (lysinopril); CalciumChannel Blockers:α[3-[[2-(3,4dimethoxyphenyl)ethyl]methylamino]-propyl]-3,4-dimethoxy-α-(1-methylethyl)benzene-acetonitrile (verapamil);1,4-dihydro-2,6-dimethyl-4-(2-nitrophenyl)-3,5-pyridinedicarboxylic aciddimethyl ester (nifedipine); 2-(2,2-dicyclohexylethyl)piperidine(perhexiline); N-(1-methyl-2-phenylethyl)- -phenylbenzenepropanamine(prenylamine);3-(aminosulfonyl)-4-chloro-N-(2,3-dihydro-2-methyl-1H-indol-1-yl)benzamide(indapamide); (2'-(2-diethylaminoethoxy)-3-phenylpropiophenone(etafenone);(4-[4,4-bis-(4-fluorophenyl)butyl]-N-(2,6-dimethylphenyl)-1-piperazineacetamide)(lidoflazine);(2-(N-benzyl-N-methylamino)ethylmethyl-2,6-dimethyl-4-(m-nitrophenyl)-1,4-dihydro-3,5-pyridinedicarboxylateHCl) (nicardipine);(N-(3,4-dimethoxyphenethyl)-2-(3,4-dimethoxyphenyl)-N-methyl-m-dithiane-2-propylamine-1,1,3,3-tetraoxide)(tiapamil);(5,6-dimethoxy-2-(3-[(α(3,4-dimethoxy)phenylethyl)-methylamino]propyl)phthalimidine)(falipamil);(β[(2-methylpropoxy)methyl]-N-phenyl-N-phenylmethyl-1-pyrrolidineethanamineHCl monohydrate) (bepridil);((±)-cis-3-(acetyloxy)-5-[2-(dimethylamino)ethyl]-2,3-dihydro-2-(4-methoxyphenyl)-1,5-benzothiazepin-4-(5H)-one)(diltiazem); (5H)-one) (diltiazem);((E)-1-[bis-(p-fluorophenyl)methyl]-4-cinnamylpiperazine di HCl)(flunarizine);(5-[(3,4-dimethoxyphenethyl)methylamino]-2-isopropyl2-(3,4,5-trimethoxyphenyl)valeronitrile(gallopamil);(ethylmethyl(2,3-dichlorophenyl)-1,4-dihydro-2,6-dimethyl-3,5-pyridinedicarboxylate(felodipine);(isopropyl-2-methoxyethyl-1,4-dihydro-2,6-dimethyl-4-(3-nitrophenyl)-3,5-pyridinecarboxylate)(nimodipine);(3-ethyl-5-methyl-1,4-dihydro-2,6-dimethyl-4-(3-nitrophenyl)-3,5-pyridine-dicarboxylate)(nitrendipine); and Other Antihypertensive Agents: aminophylline;cryptenamine acetates and tannates; deserpidine; meremethoxyllineprocaine; pargyline; tirmethaphan camsylate; and the like, as well asadmixtures and combinations thereof.
 7. A method of treatingrenin-associated hypertension or congestive heart failure in mammalscomprising administering a therapeutically-effective amount of a peptideaccording to claim
 1. 8. A method according to claim 7, wherein mammalsare human and the therapeutically-effective amount is from 0.02 to 10grams per day.
 9. A method of diagnosing renin as a contributory factorin hypertension or congestive heart failure comprising administering toa patient from 0.1 to 10 mg/kg of body weight of the patient a peptideaccording to claim 1 and monitoring the patient's blood pressure for atransitory fall that would indicate supranormal plasma renin levels.